Treatment of T-helper cell type 2-mediated immune diseases by retinoid antagonists

ABSTRACT

Retinoids with retinoid receptor antagonistic activity, pharmaceutically acceptable salts and pharmaceutically acceptable hydrolyzable esters thereof, have been found efficacious in treating T-helper cell type 2 (Th2)-mediated immune diseases, such as immunoglobulin E (IgE)-mediated allergic diseases.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a divisional of patent application Ser. No.11/131,062, filed May 17, 2005, which is a divisional of Ser. No.10/353,523, filed Jan. 29, 2003, which is a divisional of Ser. No.09/631,655, Aug. 3, 2000, now U.S. Pat. No. 610,742, which is adivisional of Ser. No. 09/189,189, Nov. 10, 1998, now U.S. Pat. No.6,133,309.

BACKGROUND OF THE INVENTION

1. Field

The present invention relates a method for using retinoid antagonists,such as retinoids with selective Retinoic Acid Receptor (“RAR”)antagonistic activity, Retinoid X Receptor (“RXR”) antagonistic activityor mixed RAR-RXR antagonistic activity, to treat T-helper cell type 2.(“Th2”)-mediated immune diseases. Th2-mediated immune diseases includeimmunoglobulin E (“IgE”)-mediated allergic diseases. The presentinvention also relates.to using retinoid antagonists to preparemedicaments for treating Th2-mediated immune diseases.

2. Description

Retinoids are a class.of compounds structurally related to vitamin A andinclude natural and synthetic compounds. Retinoids are clinically usefulin the treating dermatological and oncological diseases.

Retinoid activity is thought to be mediated by the nuclear retinoidreceptors RARα, β, γ and RXRα, β, γ, belonging to the superfamily ofsteroid, thyroid hormone vitamin D, peroxisome proliferator-activatedreceptors [Pfahl et al., Vitamins and Hormones, 49: 327-382 (1994)].Retinoids with receptor agonistic activity bind to and activatereceptors, whereas retinoids with receptor antagonistic activity bind toreceptors but do not activate them.

Experimentally, retinoids with retinoid receptor agonistic activity havebeen shown to be active in model systems for treating dermatological andoncological diseases and in models for treating immunological diseases.Retinoids with retinoid receptor agonistic activity have been shownactive in treating adjuvant arthritis [Brinckerhoffet al., Science, 221:756-758 (1983)] and experimental allergic encephalomyelitis [Massacesiet al., J. Clin. Invest. 88: 1331-1337(1991); Racke et al., J.Immunol.,154, 450-458 (1995)], animal models for rheumatoid arthritis andmultiple sclerosis, respectively. Both diseases are considered to belongto Th1-mediated immune diseases.

Experimentally, retinoids with retinoid receptor antagonistic activity(retinoid antagonists) have been shown effective in counteracting manyproperties of retinoids with retinoid receptor-agonistic activity(retinoid agonists), such as inhibiting cell proliferation, inducingcell differentiation, inducing apoptosis and inhibiting angiogenesis[Bollag et al., Int. J. Cancer, 70: 470-472 (1997)]. Retinoidantagonists also suppress toxic side effects of retinoid agonists, suchas the signs and symptoms of the hypervitaminosis A syndrome andteratogenesis [Standeven et al., Toxicol. Appl. Pharmacol., 138: 169-175(1996); Eckhardt and Schmitt, Toxicol. Letters, 70: 299-308 (1994)].Therefore, they may be useful clinically in preventing or treatingadverse events caused by retinoid agonists.

Retinoid antagonists have been proposed for clinical use in theprevention and treatment of retinoid-induced toxicity and side effects,particularly of the so-called hypervitaminosis A syndrome. Retinoidantagonists have also been proposed to be used in combination withretinoid receptor agonists or other nuclear receptor agonists forpreventing and treating preneoplastic or neoplastic lesions,vitreo-retinopathy, and retinal detachment. In addition, retinoidantagonists may be useful as single agents, based on theiranti-proliferative effect, for treatment of certain neoplasmsinsensitive to retinoid receptor agonists [WO 97/09297].

The subject invention provides for the first time a method for usingretinoid antagonists in the treatment of Th2-mediated immune diseases,such as IgE-mediated allergic diseases and diseases mediated by theTh2-related cytokines.

SUMMARY OF THE INVENTION

The subject invention provides a method of treating a T-helper cell type2-mediated immune disease or a disease mediated by T-helper cell type2-related cytokines. This method comprises administering to a subjecthaving a T-helper cell type 2-mediated immune disease or a diseasemediated by T-helper cell type 2-related cytokines an effective amountof a compound selected from the group consisting of retinoidantagonists, pharmaceutically acceptable salts of retinoid antagonists,and pharmaceutically acceptable hydrolyzable esters of such retinoidantagonists and their salts.

DETAILED DESCRIPTION OF THE INVENTION

The subject invention will now be described in terms of its preferredembodiments. These embodiments are set forth to aid in understanding theinvention but are not to be construed as limiting.

In the scope of the present invention the term “retinoid antagonists” isused for retinoids or compounds with RAR, RXR or mixed RAR-RXRantagonistic activity. It includes compounds with receptor neutralantagonistic activity (neutral antagonists), receptor inverse agonisticactivity (inverse agonists) and negative hormone activity (negativehormones) [Klein et al., J. biol Chem., 271: 22692-22696 (1996)].

In the scope of the present invention the term “retinoid antagonists”encompass compounds of formulas I-XVI depicted below:

-   a) RAR a-antagonists of formulas:    -   wherein R¹ is C₅₋₁₀-alkyl, and R² and R³ independently of each        other are hydrogen or fluorine;        such compounds are described in U.S. Pat. No. 5,391,766 and J.        Med. Chem., 40: 2445 (1997);-   b) RAR α,β antagonists of formulas:    -   wherein R⁴ is diamantyl, X is O or NH, R⁵ is phenyl or benzyl,        and    -   wherein optionally either ring A or ring B is present;        such compounds are described in Med. Chem. Res., 1: 220 (1991);        Biochem. Biophys. Res. Com., 231: 243 (1997); and J. Med. Chem.,        37: 1508 (1994);-   c) RAR β,γ antagonists of formula    -   wherein R⁶ and R⁷ independently of each other hydroxy,        C₁₋₄-alkoxy, optionally branched C₁₋₅-alkyl or adamantyl;        such compounds are described in J. Med. Chem. 38: 4993 (1995);-   d) RAR γ antagonists of formulas:    -   such compounds are described in Cancer Res., 55: 4446 (1995);-   e) RAR α, β, γ antagonists of formulas:    -   wherein Y is —CH₂— or sulfur and Z is —CH═ or nitrogen, and R⁸        is hydrogen or C₁₋₄-alkyl;        such compounds are described in J. Med. Chem. 38: 3163 and 4764        (1995); J. Biol. Chem., 271: 11897 and 22692 (1996);-   f) RXR antagonists of formulas:    -   wherein the dotted bond is optional; and, when the dotted bond        is present, R⁹ is methyl and R¹⁰ is hydrogen; and, when the        dotted bond is absent, R⁹ and R¹⁰ taken together are rethylene        to form a cis-substituted cyclopropyl ring; R¹¹ is C₁₋₄-alkoxy;        such compounds are described in EP Patent Appl. No. 97 107        843.1; J. Med. Chem., 39: 3229 (1996); and Nature, 383: 450        (1996).

In accordance with this invention, it has thus been found thatadministration of retinoid antagonists, pharmaceutically acceptablesalts, and pharmaceutically acceptable hydrolyzable esters thereof, areefficacious in treating patients with T-helper cell type 2(Th2)-mediated diseases. It has also been found that the administrationof retinoid antagonists is efficacious in treating patients withdiseases mediated by Th2-related cytokines, such as interleukin-4 (IL4)and IL-5.

The invention, therefore, in one aspect, relates to the use of retinoidantagonists, their pharmaceutically acceptable salts or pharmaceuticallyacceptable hydrolyzable esters, for the manufacture of a medicament forthe treatment of T-helper cell type 2 (Th2)-mediated immune diseases. Inanother aspect the invention relates to the use of retinoid antagonists,their pharmaceutically acceptable salts or pharmaceutically acceptableesters thereof for the manufacture of a medicament for the treatment ofdisease mediated by Th2-related cytokines, such as IL-4 and IL-5.

The invention also relates to a method for treating patients havingT-helper cell type 2 (Th2)-mediated immune diseases comprisingadministering to said human patient a compound selected from the groupof retinoid antagonists, pharmaceutically acceptable salts andpharmaceutically acceptable hydrolyzable esters thereof, said compoundbeing administered in an amount effective to treat said disease. Theterm “treatment” or “treating” includes preventive and/or therapeutictreatment.

As used herein, the term “T-helper cell type 2-mediated immune diseases”relates to diseases involving immunoglobulin E (IgE) and mast cells dueto the development and activation of allergen-specific Th2 cells and itencompasses allergic diseases, such as atopic dermatitis, otherdermatological diseases associated with atopy; allergic rhinitis or hayfever, allergic bronchial asthma in its acute or chronic, mild or severeforms, with or without acute or chronic bronchitis. Elevated serumlevels of immunoglobulin E (IgE) and hypereosinophilia can be associatedwith these diseases. Retinoid antagonists are effective in all thoseimmune diseases which are linked with an increase of Th2 cell activityand an increased secretion of the related cytokines, such as IL4 andIL-5. The therapeutic effect of retinoid antagonists is due to adecrease in Th2 cell activity, a decreased secretion of the relatedcytokines, such as IL-4 and IL-5, and/or an increase in Th1 cellactivity due to the enhancement of IL-12 production by activatedmyelomono-cytic cells. [S. Romagnani, Ann. Rev. Immunol., 12: 227-257(1994); Romagnani, ed., Th1 and Th2 Cells in Health and Disease. Chem.Immunol., Karger, Basel, 63: 187-203 (1996); Abbas et al., Nature, 383:787-793 (1996)].

The efficacy of the retinoid antagonists in accordance with the presentinvention can be shown by their ability to either upregulate Th1 cellactivity or induce/stimulate the production of cytokines, such as IL-12,IFNγ, TNF; and/or down-regulate Th2 cell activity, or inhibit theproduction of cytokines, such as IL-4 and IL-5.

Retinoid antagonists are active in the treatment of allergic bronchialasthma. The hallmarks of inflammation associated with asthmatic diseaseare the presence of activated eosinophils, an increased sensitivity ofthe airways (hyperresponsiveness), edema, mucus hypersecretion andcough. This inflammatory process is mediated by the generation andactivation of Th2-type cells. The ability of retinoid antagonists topromote a Th1-type response and thereby to suppress the Th2-typeresponse is thought to be the mechanism underlying the efficacy of thesecompounds in allergic lung inflammation/asthma. Retinoid antagonists areacting on Th1-type cells, in inhibiting the signs and symptoms ofallergic lung inflammation/asthma [Gavett et al., J. Exp. Med., 182:1527-1536 (1995); Kips et al., Am. J. Respir. Crit. Care Med., 153:535-539 (1996)]. They are active in antigen/allergen (e.g.ovalbumin)-sensitized and challenged animals. Retinoid antagonists,given either systemically or topically by aerosol, are efficacious inattenuating, inhibiting or reversing bronchoconstriction, airway edemaand mucus hypersecretion, airway inflammation, accumulation ofeosinophils and neutrophils in the broncho-alveolar tissue andbroncho-alveolar lavage respectively, as well as airwayhyperresponsiveness to non-specific stimuli.

For the treatment, the active compound, i.e. a retinoid antagonist, apharmaceutically acceptable salt or a pharmaceutically acceptablehydrolyzable ester thereof, is administered either systemically ortopically. Preferably, said compound is administered as a compositioncontaining said active compound and a pharmaceutically acceptablecarrier or diluent compatible with said active compound. In preparingsuch composition, any conventional pharmaceutically acceptable carriercan be utilized. When the drug is administered orally, it is generallyadministered at regular intervals, conveniently at mealtimes or oncedaily. It has been established that this compound is effective in doseswhich show no or only mild side effects when given orally or when giventopically. Therefore, oral or topical administration of the activecompound is generally preferred. For treating diseases of the skin,mouth, nose, pharynx, larynx, bronchus etc. oral combined with topicaladministration may also be used advantageously.

In the treatment of T-helper cell type 2-mediated immune diseases,retinoid antagonists, when administered orally do not or notsignificantly induce the adverse events belonging to the toxic syndromeof hypervitaminosis A, such as mucocutaneous, musculoskeletal,neurologic manifestations and elevation of transaminases, triglyceridesand cholesterol. In addition, they are not or less teratogenic incontrast to the receptor agonistic retinoids clinically useful in thetreatment of dermatological and oncological diseases, such as all-transretinoic acid (tretinoin), 13-cis retinoic acid (isotretinoin),etretinate and acitretin.

In the treatment of T-helper cell type 2-mediated immune diseases,retinoid antagonists, pharmaceutically acceptable salts orpharmaceutically acceptable hydrolyzable esters thereof, can be usedalone or in combination with other measures, e.g. in combination withother pharmaceutically active substances such as topical or systemiccorticosteroids, antihistaminics and bronchodilating agents. If used incombination with other substances, retinoid antagonists and said othersubstances can be administered separately or incorporated in effectiveamounts into one pharmaceutical composition.

In the scope of the present invention, the “pharmaceutically acceptablesalts” includes any salt chemically permissible in the art for retinoidantagonists and applicable to human patients in a pharmaceuticallyacceptable preparation. Any such conventional pharmaceuticallyacceptable salt of retinoid antagonists can be utilized. Among theconventional salts which can be utilized, there are the base saltsincluded, for example, alkali metal salts such as the sodium orpotassium salt, alkaline earth metal salts such as the calcium ormagnesium salt, and ammonium or alkyl ammonium salts.

In accordance with this invention the retinoid antagonists can also beadministered in the form of its pharmaceutically acceptable hydrolyzableesters. Any pharmaceutically acceptable hydrolyzable ester can be usedin the compositions and methods of this invention. Among the preferredesters are: the aromatic esters such as benzyl esters in which thebenzyl moiety is unsubstituted or substituted with lower alkyl, halo,nitro, thio, or substituted thio; or lower alkyl esters, e.g. ethyl,t-butyl, cyclopentyl, cyclohexyl or cycloheptyl ester; or.9-fluorenylmethyl ester.

In the scope of the present invention the term “alkyl” meansstraight-chain, branched or cyclic alkyl residues, in particular thosecontaining from 1 to 12 carbon atoms, such as methyl, ethyl, propyl,isopropyl, t-butyl, decyl, dodecyl, cyclopentyl, cyclohexyl, cycloheptyland the like. The term “lower alkyl” means alkyl groups containing from1 to 7 carbon atoms.

The aforementioned retinoid antagonists, the salts and esters thereofare useful especially in pharmaceutically acceptable oral or topicalmodes. These pharmaceutical compositions contain said active compound inassociation with a compatible pharmaceutically acceptable carriermaterial. Any conventional carrier material can be utilized. The carriermaterial can be organic or inorganic inert carrier material suitable fororal administration. Suitable carriers include water, gelatin, gumarabic, lactose, starch, magnesium stearate, talc, vegetable oils,polyalkylene-glycols, petroleum jelly and the like. Furthermore, thepharmaceutically active preparations may contain other pharmaceuticallyactive agents. Additionally, additives such as flavoring agents,preservatives, stabilizers, emulsifying agents, buffers and the like maybe added in accordance with accepted practices of pharmaceuticalcompounding.

The pharmaceutical preparations can be made up in any conventional formincluding: (a) solid form for oral administration such as tablets,capsules (e.g. hard or soft gelatine capsules), pills, sachets, powders,granules, and the like; and (b) preparations for topical administrationssuch as solutions, suspensions, ointments, creams, gels, micronizedpowders, sprays, aerosols and the like. Pharmaceutical preparations maybe sterilized and/or may contain adjuvants such as preservatives,stabilizers, wetting agents, emulsifiers, salts for varying the osmoticpressure and/or buffers.

For topical administration to the skin or mucous membrane theaforementioned derivative is preferably prepared as ointments,tinctures, creams, gels, solution, lotions, sprays; aerosols and drypowder for inhalation, suspensions, shampoos, hair soaps, perfumes andthe like. In fact, any conventional composition can be utilized in thisinvention. Among the preferred methods of applying the compositioncontaining the agents of this invention is in the form of an ointment,gel, cream, lotion, spray; aerosol or dry powder for inhalation. Apharmaceutical preparation for topical administration to the skin can beprepared by mixing the aforementioned active ingredient with non-toxic,therapeutically inert, solid or liquid carriers customarily used in suchpreparation. These preparations generally contain 0.01 to 5.0 percent byweight, preferably 0.1 to 1.0 percent by weight, of the activeingredient, based on the total weight of the composition.

In preparing the topical preparations described above, additives such aspreservatives, thickeners, perfumes and the like conventional in the artof pharmaceutical compounding of topical preparation can be used. Inaddition, conventional antioxidants or mixtures of conventionalantioxidants can be incorporated into the topical preparationscontaining the afore-mentioned active agent. Among the conventionalantioxidants which can be utilized in these preparations are includedN-methyl-α-tocopherolamine, tocopherols, butylated hydroxyanisole,butylated hydroxytoluene, ethoxyquin and the like. Cream-basepharmaceutical formulations containing the active agent, used inaccordance with this invention, are composed of aqueous emulsionscontaining a fatty acid alcohol, semi-solid petroleum hydrocarbon,ethylene glycol and an emulsifying agent.

Ointment formulations containing the active agent in accordance withthis invention comprise admixtures of a semi-solid petroleum hydrocarbonwith a solvent dispersion of the active material. Cream compositionscontaining the active ingredient for use in this invention preferablycomprise emulsions formed from a water phase of a humectant, a viscositystabilizer and water, an oil phase of a fatty acid alcohol, a semi-solidpetroleum hydrocarbon and an emulsifying agent and a phase containingthe active agent dispersed in an aqueous stabilizer-buffer solution.Stabilizers may be added to the topical preparation. Any conventionalstabilizer can be utilized in accordance with this invention. In the oilphase, fatty acid alcohol components function as a stabilizer. Thesefatty acid alcohol components function as a stabilizer. These fatty acidalcohol components are derived from the reduction of a long-chainsaturated fatty acid containing at least 14 carbon atoms. Also,conventional perfumes and lotions generally utilized in topicalpreparation for the hair can be utilized in accordance with thisinvention. Furthermore, if desired, conventional emulsifying agents canbe utilized in the topical preparations of this invention.

For topical treatment of allergic rhinitis and allergic bronchial asthmanasal and inhalation aerosols are used. Formulations for such aerosolsare described in Drugs and Pharmaceutical Sciences, Marcel Dekker, NewYork, 72: 547-574 (1996). Furthermore, the active compound can bedelivered by dry powder inhalation. Such formulations and devices aredescribed in Pharmaceutical Technology, June 1997, pp. 117-125.

A preferred oral dosage form comprises tablets, pills, sachets, orcapsules of hard or soft gelatin, methylcellulose or of another suitablematerial easily dissolved in the digestive tract. Each tablet, pill,sachet or capsule can preferably contain from about 5 to about 200 mg,more preferably from about 20 to about 100 mg, of active ingredient. Theoral dosages contemplated in accordance with the present invention willvary in accordance with the needs of the individual patient asdetermined by the prescribing physician. Generally, however, adaily-dosage of from 0.05 to 20 mg per kg of body weight, preferably 0.1to 7 mg, and most preferably from about 0.3 mg to about 1.5 mg per kg ofbody weight of the patient is utilized. This-dosage may be administeredaccording to any dosage schedule determined by the physician inaccordance with the requirements of the patient.

The dosage for treatment typically depends on the route ofadministration, the age, weight and disease condition of the individual.Suitable dosage forms are known in the art or can be easily obtained ina manner known per se. Formulations of lotions, gels, creams, sprays;aerosols and dry powder for inhalation, hard or soft gelatin capsules,tablets and sachets that are particularly suitable in the scope of thepresent invention or that can be easily adjusted in accordance with theabove teaching are in the art.

Experimental Methods

I. In Vitro Assay for IL-12 Induction by Retinoid Antagonists

THP-1 cells were obtained from American Tissue Culture Collection andcultured in complete medium. To assay for IL-12 production, THP-1 cells,1.25×10⁶ cells/ml, were stimulated with S. aureus Cowan strain (SAC)(1/1000) and human recombinant interferon-γ (hulFN-γ) (1000 U/ml) [Ma etal., J. Exp. Med. 183: 147-157 (1996)]. Alternatively, 0.5×10⁶ humanperipheral blood mononuclear cells (PBMC) (1 ml culture in 48 wellplates) were primed with hulFN-γ (1000 U/ml) for 16 hours at 37° C., andthen stimulated with SAC (1/1000). Supernatants were collected after 48hours, and freezed at −20° C. until assayed [Panina-Bordignon et al., J.Clin. Invest. 100: 1513-1519 (1997)].

IL-12 production was measured by specific enzyme linked immunosorbantassay (ELISA), using 20C2 antibody (rat anti human IL-12 heterodimerp40-p35), at 2.5 μg/ml in coating buffer, and peroxidase-conjugated 4D6antibody (rat anti human IL-12) at 250 ng/ml in assay buffer asdescribed [Zhang et al., J. Clin. Invest. 93: 1733-1739 (1994)].Standard (recombinant human IL-12, 800 pg/ml to 6 pg/ml) and samples(100 μl) diluted in assay buffer were added to duplicate wells.Absorbance was read at 450-650 nm. The unknown IL-12 concentrations ofthe samples were read from the corresponding standard curve andmultiplied by the corresponding dilution factor. Maximal IL-12production varied between 200 and 400 pg/ml.

Lyophilized retinoid antagonists were diluted in DMSO under yellowlight, on ice at a concentration of 2 mM. Serial dilutions (1 μM-1 pM)were prepared in complete RPMI medium. 10 μl of each dilution was addedto 1 ml culture.

The results of the experiments indicate that the tested retinoidantagonists influence IL-12 production. In particular, the testedretinoid antagonists stimulate IL-12 production by activated humanmonocytes, see Table I and II. TABLE I Retinoid antagonists specificallyenhance IL-12 production by activated monocytes IL-12 IL-10 TNF-α nM(pg/ml) (pg/ml) (pg/ml) medium 0 <10 <10 SAC + IFN-γ 120 1040 1840 RAR αantagonist 1000 251 1343 1912 Compound A 100 102 1050 1600 10 n.d. 10601392 medium 0 <10 <10 SAC + IFN-γ 126 1040 2000 RAR αβγ antagonist 1000321 1116 2884 Compound B 100 205 983 2752 10 173 971 2592 medium 0 <10<10 SAC + IFN-γ 120 1040 1840 RXR antagonist 1000 298 1700 1560 CompoundC 100 161 1521 1812 10 106 1020 1484

TABLE II Retinoid antagonists enhance IL-12 production by PBMC and THP-1cells that have been primed with IFNγ and stimulated with SAC Com-Receptor Time * PBMC THP-1 pound Specificity Activity Stimuli (hrs)IL-12 (pg/ml) A RARα antagonist IFNγ + 0 503 306 SAC 16 401 nd B RARα,β, γ antagonist IFNγ + 0 371 364 SAC 16 367 nd C RXR antagonist IFNγ + 0568 577 SAC 16 367 nd none none <12 <2 IFNγ + 360 275 SAC* retinoid antagonists (1 μg) were added at time 0 together with IFNγ orafter 16 hours together with SAC.Compound Ap-[(E)-2-[3′,4′-Dihydro-4′,4′-dimethyl-7′-(heptyloxy)-2′H-1-benzothio-pyran-6′-yl]propenyl]benzoicacid 1′,1′-dioxideCompound B4-(7,7,10,10-Tetramethyl-1-pyridin-3-ylmethyl-4,5,7,8,9,10-hexahydro-1H-naphto[2,3-g]indol-3-yl)-benzoicacidCompound C(2E,4E,6Z)-7-[2-Butoxy-3,5-bis(1,1-dimethylethyl)phenyl]-3-methyl-2,4,6-octatrienoicacidII. In vitro assay for inhibition of differentiation of human naive Tcells into T helper 2 (Th2) cells by retinoid antagonists.

Naive T cells from cord blood were isolated and treated as described.[Panina-Bordignon et al. J. Clin. Invest. 100: 1513-1519 (1997)].Briefly, cord blood derived mononuclear cells were incubated withanti-CD45RA and anti-CD4 monoclonal antibodies. After a 20 minuteincubation, cells were washed and incubated with goat anti-mouseIg-coated magnetic beads. Positive cells were separated and seeded at1×10⁶ cells/ml in a 24 well plate, together with autologous adherentcells, PHA, and IL4 in the presence or absence ofp-[(E)-2-[3′,4′-Dihydro-4′,4′-dimethyl-7′-(heptyloxy)-2′H-1-benzothiopyran-6′-yl]propenyl]benzoicacid 1′,1′-dioxide (Compound A) or(2E,4E,6Z)-7-[2-Butoxy-3,5-bis(1,1-dimethylethyl)phenyl]-3-methyl-2,4,6-octatrienoicacid (Compound C) at 1 mM for 5 days. Cells were then washed and putback in culture in the presence of IL-2 (100 U/ml). After 10 days, thecells were collected and restimulated with PMA (50 ng/ml) and ionomycin(1 μg/ml) for 4 hours. Brefeldin A (10 μg/ml) was added for the last 2hours. Then the cells were fixed with 4% paraformaldehyde andpermeabilized with saponin. Fixed cells were stained with FITC-anti IFNγand PE-anti-IL-4mAbs and subjected to cytofluorimetric analysis.

The results of the experiment indicate that the tested retinoidantagonists reduce the differentiation of naive T cells intoIL-4-secreting Th2 cells. (Table III) TABLE III Suppression of IL-4expression in Th2 cells by retinoid antagonists IL-4 expressing cells %gated cells % Th2 cells Th2 26.32 100 Th2 + Compound A 10.8 41 Th2 +Compound C 8.5 32III. Murine Model of Allergen-Induced Airway Inflammation andHyperresponsiveness.

C57BL/6 mice (8-9 weeks old). are actively sensitized to ovalbumin (OA)on day 0 and on day 14 by a intraperitoneal injection of 10 μg OA+1 mgAl(OH)₃ (gel suspension) in 0.2 ml sterile saline. On day 21, the micewere challenged with 5.0% OA aerosol for 18 minutes. The aerosol isgenerated by a De Vilbiss Ultra-Neb 90 ultrasonic nebulizer, the outletof which is connected to a small plexiglass chamber containing theanimals. The mice are dosed with the RXR antagonist Compound C (10 and30 mg/kg intraperitoneally) daily for three days, 48 hours, 24 hours,and immediately prior. to OA challenge. Animals are used on day 21.

Airway Inflammatory Cell Accumulation

On day 24, three days after the challenge with OA aerosol, animals areanesthetized with urethane (2.4<g/kg) and tracheotomized with a 23 gaugecatheter. Lungs are lavaged with aliquots (2×1 ml) of sterile Hank'sbalanced salt solution without Ca⁺⁺ and Mg⁺⁺. Lavage fluid is recoveredafter 30 sec. by gentle aspiration and pooled for each animal. Samplesthen are centrifuged at 2000 rpm for 15 minutes at. 5° C. Red bloodcells are lysed from the resulting pellet with 0.5 ml distilled waterand the cells remaining in the pellet are reconstituted with 5 ml HBSS.Samples are centrifuged a second time at 2000 rpm for 15 minutes at 5°C. The resulting pellet is suspended in 1 ml of HBSS. Total cell numberis determined from an aliquot of the cell suspension using ahemocytometer. For cytological preparations, the cells are fixed oncytocentrifuged slides stained with a modified Wright's stain.Differential counts on at least 300 cells are made using standardmorphological criteria to classify cells.

The results of the experiments indicate that the tested retinoidantagonists inhibit the allergen-induced accumulation of airwayinflammatory cells (Table IV). TABLE IV Suppression of airwayinflammatory cell accumulation by retinoid antagonists in a mouse modelof allergen-induced airway inflammation Cell Influx Percent of(cells/ml) reduction Compound C Compound C Vehicle 10 mg/kg 30 mg/kg 10mg/kg 30 mg/kg Total 795000 488000 271000 39% 66% leukocytes Macrophages443000 289000 172000 35% 62% Eosinophils 335000 176000 91000 48% 73%Airway Hyperresponsiveness

On day 24, three days after the challenge with OA aerosol, animals areanesthetized with pentobarbital sodium (100 mg/kg, i.p.) andtracheotomized (PE-190). A jugular vein is cannulated with a sylastictubing for i.v. drug delivery. Animals are placed in a whole bodyplethysmograph with a built-in pneumotachograph and mechanicallyventilated (V_(f)=150/min., V_(t)=0.3 ml; Model 683, Harvard Apparatus,S. Natic, Mass.) immediately following pancuronium bromide (0.1 mg/kg,i.v.) treatment. Tidal volume is obtained from an integration of therespiratory flow signal using a differential pressure transducer(Validyne DP 103-08, Northridge, Calif.). Transpulmonary pressure ismeasured with a differential pressure transducer (Validyne DP 45-30,Northridge. Calif.) as the difference between intratracheal pressure andintrapleural pressure (obtained from a cannula inserted into theintercostal space). Changes in lung resistance (cm H₂O/Ml/s) toincreasing doses of methacholine (30, 100, 300, 1000 μg/kg, i.v.) arecalculated from transpulmonary pressure, tidal volume, and respiratoryflow measurements using a Modular Instrument Signal Processing System(Malvern, Pa.).

The results of the experiments indicate that retinoid antagonists canprevent or reverse allergic airway inflammation and inhibitantigen-induced bronchoconstriction, typical for allergic airwaydiseases, such as allergic bronchial asthma.

Examples for formulations: capsules, tablets, sachets, lotions, gels,creams, aerosols and dry powder for inhalation. The active compounds inthe following examples are

-   p-[(E)-2-[3′, 4′-Dihydro-4,4′-dimethyl-7′-(heptyloxy)-2′H-1    -benzothio-pyran-6′-yl]propenyl]benzoic acid 1,1′-dioxide or-   (2E,4E,6Z)-7-[2-Butoxy-3,5-bis(1,1-dimethylethyl)phenyl]-3-methyl-2,4,6-octatrienoic    acid.

EXAMPLE 1

Lotion (solution) preferred Active compound 0.1-2.0 g Propylene Glycol5.00-20.00 g 10.00 g PEG-Glyceryl Cocoate * 0.00-20.00 g 10.00 gdl-a-Tocopherol 0.001-0.50 g 0.02 g Ascorbyl Palmitate 0.01-0.20 g 0.10g Propyl Gallate 0.001-0.02 g 0.002 g Citric acid, anhydr. ** 0.00-0.20g 0.01 g Isopropanol *** 40.00-90.00 g 50.00 g Water, dem. ad 100.00 g100.00 g resp. ml* or other tensides** or other complexing agents e.g. EDTA*** or other alcohols e.g. Ethanol

EXAMPLE 2

Gel preferred Active compound 0.1-2.0 g Propylene Glycol 5.00-20.00 g10.00 g PEG-Glyceryl Cocoate * 0.00-20.00 g 10.00 g dl-α-Tocopherol0.001-0.50 g 0.02 g Ascorbyl Palmitate 0.01-0.20 g 0.10 g Propyl Gallate0.001-0.02 g 0.002 g Citric acid, anhydr. ** 0.00-0.20 g 0.01 gIsopropanol *** 40.00-90.00 g 50.00 g HPMC **** 0.50-5.00 g 3.00 gPreservative ***** q.s. q.s. Water, dem. ad 100.00 g 100.00 g resp. ml* or other tensides** or other complexing agents e.g. EDTA*** or other alcohols e.g. Ethanol**** Hydroxypropyl Methylcellulose or other polymers e.g. neutralisedCarbomer, Methyl Cellulose, Sodium Carboxymethylcellulose***** Preservatives e.g., Paraben esters (methyl, ethyl, propyl, butyl).Sorbic Acid. Benzoic Acid

EXAMPLE 3

Cream preferred Active compound 0.1-2.0 g Glycerol 0.00-10.00 g 5.00 gNa₂ EDTA 0.001-0.50 g 0.03 g Glycerides * 5.00-20.00 g 10.00 g CetylAlcohol 0.50-5.00 g 1.00 g Stearyl Alcohol 0.50-5.00 g 1.00 g Glycerolmono Stearate 1.00-8.00 g 4.00 g Ceteareth ** 0.50-5.00 g 2.00 gdl-α-Tocopherol 0.001-0.50 g 0.02 g Preservative *** q.s. q.s. Water,dem. ad 100.00 g 100.00 g* e.g. Caprylic/Capric/Triglyceride, Caprylic/Capric/LinoleicTriglycerides, natural glycerides, as well as e.g. Propylene Glycol,Dicaprylate/Dicaprate and waxes, such as Stearyl, Stearate, OleylOleate, Isopropyl Myristate** Ceteareth 5-30, or other emulsifiers such as Polysorbase 20-80,Sorbitane esters of fatty acids, fatty acid esters of PEG.*** Preservatives e.g., Paraben esters (methyl, ethyl, propyl, butyl).Sorbic Acid. Benzoic Acid

EXAMPLE 4

Fill mass for soft gelatin capsules Active compound 5.0-200.0 mg Oil *1-3 parts Wax mixture ** 1-5 parts Fill volume 1-6 minims 20 mg SoftGelatin Capsules Ingredients mg/capsule Active compound 20.000dl-α-Tocopherol 0.028 Hydrogenated Castor Oil 4.200Caprylic/Capric/Stearic Triglyceride 56.000 (Synthetic Triglyceride)Triglyceride, Medium Chain 199.772 Total 280.000 mg* natural vegetable oils, e.g. soy oil, peanut oil, and artificialglycerides** composition of natural and artificial waxes or partially hydratedfats

EXAMPLE 5

Hard Gelatine capsules containing 20 mg active substance: Composition:One Capsule contains: Active compound 20.0 mg Gelatine Bloom 30 70.0 mgMaltodextrin MD 05 108.0 mg dl-α-Tocopherol 2.0 mg Sodium ascorbate 10.0mg Microcrystalline cellulose 48.0 mg Magnesium stearate 2.0 mg (weightcapsule content) 260.0 mgProcedure:

The active substance is wet milled in a solution of gelatine,maltodextrin, dl-α-Tocopherol and sodium ascorbate.

The wet milled suspension is spray-dried.

The spray-dried powder is mixed with microcrystalline cellulose andmagnesium stearate.

260 mg each of this mixture are filled into hard gelatine capsules ofsuitable size and color.

EXAMPLE 6

Tablet containing 20 mg active substance: Composition: Tablet kernel:Active compound 20.0 mg Anhydrous lactose 130.5 mg MicrocrystallineCellulose 80.0 mg dl-α-Tocopherol 2.0 mg Sodium ascorbate 10.0 mgPolyvinylpyrrolidone K30 5.0 mg Magnesium stearate 2.5 mg (Kernelweight) 250.0 mg Film coat: Hydroxypropyl methylcellulose 3.5 mgPolyethylenglycol 6000 0.8 mg Talc 1.3 mg Irone oxide, yellow 0.8 mgTitanium dioxide 0.8 mg (weight of the film) 7.4 mgProcedure:

The compound is mixed with anhydrous lactose and microcrystallinecellulose. The mixture is granulated in water with a solution/dispersionof polyvinylpyrrolidone, dl-α-Tocopherol and sodium ascorbate.

The granular material is mixed with magnesium stearate and afterwardspressed as kernels with 250 mg weight.

The kernels are film coated with a solution/suspension ofabove-mentioned compositions.

EXAMPLE 7

Sachet containing active substance Composition: Active compound 200.0 mgLactose, fine powder 990.0 mg Microcrystalline Cellulose 1250.0 mgSodium Carboxymethyl cellulose 14.0 mg dl-α-Tocopherol 5.0 mg Sodiumascorbate 20.0 mg Polyvinylpyrrolidone K30 10.0 mg Magnesium stearate10.0 mg

EXAMPLE 8

Aerosol for inhalation, metered dose inhaler Active compound 0.5%(0.1-2.0%) Sorbitantrioleate 5% dl-α-Tocopherol 0.4% Propellant (mixtureof Trichlorofluoro- 94.1% methane and Dichlorodifluoromethane)

EXAMPLE 9

Dry powder inhaler Active compound * 0.5 mg (0.1 mg-2.0 mg) Lactosemonohydrate  25 mg* jet-milled, spray-dried

1. A method of treating a T-helper cell type 2-mediated immune diseaseor a disease mediated by T-helper cell type 2-related cytokines, whichcomprises administering to a subject having a T-helper cell type2-mediated immune disease or a disease mediated by T-helper cell type2-related cytokines an effective amount of a compound selected from thegroup consisting of retinoid antagonists, pharmaceutically acceptablesalts of retinoid antagonists,and pharmaceutically acceptablehydrolyzable esters of such retinoid antagonists and their salts.
 2. Themethod of claim 1, wherein the T-helper cell type 2-mediated immunedisease is an immunoglobulin E-mediated allergic disease.
 3. The methodof claim 2, wherein the immunoglobulin E-mediated allergic disease isatopic dermatitis.
 4. The method of claim 2, wherein the immunoglobulinE-mediated allergic disease is allergic rhinitis.
 5. The method of claim2, wherein the immunoglobulin E-mediated allergic disease is allergicbronchial asthma.
 6. The method of claim 1, wherein the disease mediatedby Th2-related cytokines is mediated by IL-4 or IL-5.
 7. The method ofclaim 1, wherein the compound is a retinoid antagonist of the formula:

wherein R¹ is C₅₋₁₀-alkyl, or a pharmaceutically acceptable salt of suchretinoid antagonist or a pharmaceutically acceptable hydrolyzable esterof such retinoid antagonist or its salt.
 8. The method of claim 1,wherein the compound is a retinoid antagonist of the formula:

wherein R¹ is C₅₋₁₀-alkyl, or a pharmaceutically acceptable salt of suchretinoid antagonist or a pharmaceutically acceptable hydrolyzable esterof such retinoid antagonist or its salt.
 9. The method of claim 1,wherein the compound is a retinoid antagonist of the formula:

wherein R² and R³ are each independently hydrogen or fluorine, or apharmaceutically acceptable salt of such retinoid antagonist or apharmaceutically acceptable hydrolyzable ester of such retinoidantagonist or its salt.
 10. The method of claim 1, wherein the compoundis a retinoid antagonist of the formula:

wherein R² and R³ are each independently hydrogen or fluorine, or apharmaceutically acceptable salt of such retinoid antagonist or apharmaceutically acceptable hydrolyzable ester of such retinoidantagonist or its salt.
 11. The method of claim 1, wherein the compoundis a retinoid antagonist of the formula:

wherein R⁴ is diamantyl, X is O or NH, or a pharmaceutically acceptablesalt of such retinoid antagonist or a pharmaceutically acceptablehydrolyzable ester of such retinoid antagonist or its salt.
 12. Themethod of claim 1, wherein the compound is a retinoid antagonist of theformula:

wherein either ring A or ring B is present, or a pharmaceuticallyacceptable salt of such retinoid antagonist or a pharmaceuticallyacceptable hydrolyzable ester of such retinoid antagonist or its salt.13. The method of claim 1, wherein the compound is a retinoid antagonistof the formula:

wherein R⁵ is phenyl or benzyl, or a pharmaceutically acceptable salt ofsuch retinoid antagonist or a pharmaceutically acceptable hydrolyzableester of such retinoid antagonist or its salt.
 14. The method of claim1, wherein the compound is a retinoid antagonist of the formula:

wherein R⁶ and R⁷ are each independently hydroxy, C₁₋₄-alkoxy,C₁₋₅-alkyl, branched C₁₋₅-alkyl, or adamantyl, or a pharmaceuticallyacceptable salt of such retinoid antagonist or a pharmaceuticallyacceptable hydrolyzable ester of such retinoid antagonist or its salt.15. The method of claim 1, wherein the compound is a retinoid antagonistof the formula:

or a pharmaceutically acceptable salt of such retinoid antagonist or apharmaceutically acceptable hydrolyzable ester of such retinoidantagonist or its salt.
 16. The method of claim 1, wherein the compoundis a retinoid antagonist of the formula:

or a pharmaceutically acceptable salt of such retinoid antagonist or apharmaceutically acceptable hydrolyzable ester of such retinoidantagonist or its salt.
 17. The method of claim 1, wherein the compoundis a retinoid antagonist of the formula:

wherein Y is —CH₂— or sulfur and Z is —CH═or nitrogen, or apharmaceutically acceptable salt of such retinoid antagonist or apharmaceutically acceptable hydrolyzable ester of such retinoidantagonist or its salt.
 18. The method of claim 1, wherein the compoundis a retinoid antagonist of the formula:

or a pharmaceutically acceptable salt of such retinoid antagonist or apharmaceutically acceptable hydrolyzable ester of such retinoidantagonist or its salt.
 19. The method of claim 1, wherein the compoundis a retinoid antagonist of the formula:

R⁸ is hydrogen or C₁₋₄-alkyl; or a pharmaceutically acceptable salt ofsuch retinoid antagonist or a pharmaceutically acceptable hydrolyzableester of such retinoid antagonist or its salt.
 20. The method of claim1, wherein the compound is a retinoid antagonist of the formula:

or a pharmaceutically acceptable salt of such retinoid antagonist or apharmaceutically acceptable hydrolyzable ester of such retinoidantagonist or its salt.
 21. The method of claim 1, wherein the compoundis a retinoid antagonist of the formula:

wherein

is a single bond or a double bond, when a double bond is present, R⁹ ismethyl and R¹⁰ is hydrogen, and when a single bond is present, R⁹ andR¹⁰ taken together are methylene thus forming a cis-substitutedcyclopropyl ring, and R¹¹ is C₁₋₄-alkoxy, or a pharmaceuticallyacceptable salt of such retinoid antagonist or a pharmaceuticallyacceptable hydrolyzable ester of such retinoid antagonist or its salt.22. The method of claim 1, wherein the compound is a retinoid antagonistof the formula:

wherein R¹¹ is C₁₋₄-alkoxy, or a pharmaceutically acceptable salt ofsuch retinoid antagonist or a pharmaceutically acceptable hydrolyzableester of such retinoid antagonist or its salt.
 23. The method of claim1, wherein the administering comprises oral administration.
 24. Themethod of claim 23, wherein the oral administration is at a daily dosageof from about 0.05 mg to about 20 mg of the compound per kg of bodyweight of the subject.
 25. The method of claim 24, wherein the oraladministration is at a daily dosage of from about 0.3 mg to about 1.5 mgof the compound per kg of body weight of the subject.
 26. The method ofclaim 23, wherein the oral administration comprises administering atablet, capsule, pill or sachet containing from about 5 mg to about 200mg of the compound.
 27. The method of claim 26, wherein the oraladministration comprises administering a tablet, capsule, pill or sachetcontaining from about 20 mg to about 100 mg of the compound.
 28. Themethod of claim 1, wherein the administering comprises topicaladministration.
 29. The method of claim 28, wherein the topicaladministration comprises administering an ointment, cream, lotion, orspray containing from about 0.01 percent to about 5.0 percent by weightof the compound.
 30. The method of claim 29, wherein the topicaladministration comprises administering an ointment, cream, lotion, orspray containing from about 0.1 percent to about 1.0 percent by weightof the compound.
 31. The method of claim 1, wherein the administeringcomprises inhalation.
 32. The method of claim 31, wherein the inhalationcomprises administering a nasal aerosol, aerosol for inhalation, or drypowder for inhalation containing from about 0.01 percent to about 5.0percent by weight of the compound.
 33. The method of claim 32, whereinthe inhalation comprises administering a nasal aerosol, aerosol forinhalations, or dry powder for inhalation containing from about 0.1percent to about 1.0 percent by weight of the compound.
 34. The methodof claim 1, wherein the compound is a retinoid antagonists or a alkalimetal salt, alkaline earth metal salt, benzyl ester, lower alkyl ester,or 9-fluorenylmethyl ester thereof.